We seek to understand how neurotransmitters signal through heterotrimeric G proteins to modulate the activities of neurons. We will focus on the mechanism of signaling by serotonin, defects in which are thought to underlie major depression in humans. Our goal is to understand how the many G protein coupled serotonin receptors function together to mediate appropriate responses to this neurotransmitter, and to find out what happens downstream of receptor activation. C. elegans uses serotonin as a neurotransmitter, and we will use this model system to investigate the mechanism of three different serotonin signaling events. In our first aim, we will characterize a signaling complex that mediates serotonin signaling onto the C. elegans egg-laying muscles. We will test a model in which multiple serotonin receptors and an ERG potassium channel are co-localized to postsynaptic structures so that localized signals produced by serotonin receptor signaling can inhibit the ERG channel to promote muscle contraction. In the second aim, we will study the mechanism by which serotonin released onto the egg-laying muscles also signals back onto the neurons that release it via autoreceptors to feedback inhibit further serotonin release. We will use a combination of genetic and biochemical studies to analyze two new types of signaling regulators that our preliminary results show enhance such feedback signaling: a GPR/GoLoco protein which binds G1 subunits, and the RIC-8 protein which acts as a G protein nucleotide exchange factor. In the third aim, we will study the mechanism by which serotonin signals to slow down C. elegans locomotion behavior. We have used a genetic screen to identify genes required for this serotonin signaling event. We found that two different serotonin receptors are each required for the effect of serotonin on locomotion: knocking out either receptor alone or both together leads to the same strong defect in serotonin response. We will study how these two different serotonin receptors work together. We will also comprehensively analyze a new protein identified by our screen that is required specifically for serotonin signaling, but that is not similar to any protein previously known to be involved in serotonin signaling. The mechanisms of serotonin signaling are highly conserved between C. elegans and humans and the insights made possible by the power of the C. elegans model system should shed light on new details of human serotonin signaling. This will ultimately help to understand the causes of depression and the interventions that can be used to treat it. PUBLIC HEALTH RELEVANCE: This proposal investigates the basic mechanisms by which neurons in the brain communicate with each other using chemical neurotransmitters, focusing in particular on the neurotransmitter serotonin. Defects in signaling by serotonin and other neurotransmitters appear to underlie depression, schizophrenia and other disorders of mental health, but the precise details of these defects are in most cases unknown. Understanding in detail how neurotransmitters act should make it possible to precisely define the defects that cause mental disorders and to thus develop effective treatments for them.)